title: Fusion of lysosomes with late endosomes produces a hybrid organelle of intermediate density and is NSF dependent.
abstract: Using a cell-free content mixing assay containing rat liver endosomes and lysosomes in the presence of pig brain cytosol, we demonstrated that after incubation at 37 degrees C, late endosome-lysosome hybrid organelles were formed, which could be isolated by density gradient centrifugation. ImmunoEM showed that the hybrids contained both an endocytosed marker and a lysosomal enzyme. Formation of the hybrid organelles appeared not to require vesicular transport between late endosomes and lysosomes but occurred as a result of direct fusion. Hybrid organelles with similar properties were isolated directly from rat liver homogenates and thus were not an artifact of cell-free incubations. Direct fusion between late endosomes and lysosomes was an N-ethylmaleimide-sensitive factor-dependent event and was inhibited by GDP-dissociation inhibitor, indicating a requirement for a rab protein. We suggest that in cells, delivery of endocytosed ligands to an organelle where proteolytic digestion occurs is mediated by direct fusion of late endosomes with lysosomes. The consequences of this fusion to the maintenance and function of lysosomes are discussed.
authors.affiliation: Department of Clinical Biochemistry, University of Cambridge, Addenbrooke's Hospital, Cambridge, CB2 2QR, United Kingdom.
authors.forename: B M
authors.initials: BM
authors.lastname: Mullock
authors.forename: N A
authors.initials: NA
authors.lastname: Bright
authors.forename: C W
authors.initials: CW
authors.lastname: Fearon
authors.forename: S R
authors.initials: SR
authors.lastname: Gray
authors.forename: J P
authors.initials: JP
authors.lastname: Luzio
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002352:Carrier Proteins / Q000494:pharmacology*; D002474:Cell-Free System; D002499:Centrifugation, Density Gradient; D002867:Chromones / Q000494:pharmacology; D003600:Cytosol / Q000502:physiology; D004705:Endocytosis; D011992:Endosomes / Q000502:physiology* / Q000648:ultrastructure; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000502:physiology; D020730:Guanine Nucleotide Dissociation Inhibitors; D006160:Guanosine Triphosphate / Q000378:metabolism; D008099:Liver / Q000648:ultrastructure; D008247:Lysosomes / Q000502:physiology* / Q000648:ultrastructure; D008561:Membrane Fusion; D008565:Membrane Proteins / Q000494:pharmacology; D009025:Morpholines / Q000494:pharmacology; D052066:N-Ethylmaleimide-Sensitive Proteins; D000081082:Phosphoinositide-3 Kinase Inhibitors; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D013552:Swine; D033921:Vesicular Transport Proteins; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D002352:Carrier Proteins; D002867:Chromones; D004791:Enzyme Inhibitors; C086796:GDP dissociation inhibitor 1; D020730:Guanine Nucleotide Dissociation Inhibitors; D008565:Membrane Proteins; D009025:Morpholines; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011994:Recombinant Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D006160:Guanosine Triphosphate; D000255:Adenosine Triphosphate; D019204:GTP-Binding Proteins; D052066:N-Ethylmaleimide-Sensitive Proteins; C498737:Nsf protein, rat; D005033:Ethylmaleimide; D000077191:Wortmannin